Review Article

Fungal Alterations in Inflammatory Bowel Diseases

Siu Lam; Tao Zuo; Martin Ho; Francis K. L. Chan; Paul K. S. Chan; Siew C. Ng

Disclosures

Aliment Pharmacol Ther. 2019;50(11):1159-1171. 

In This Article

Advances in Mycobiota Profiling in Humans

The advent of more affordable, high-throughput sequencing technologies has improved our understanding of the fungal composition in humans. Five commercial extraction kits are frequently used for DNA extraction: QIAamp Fast DNA Stool Mini Kit (Q), Q and Bead beating, Q and Lyticase lysis buffer, FastDNA® SPIN Kit and Repeat bead beating plus column (RBBC).[24] Q gives a relatively low DNA yield but higher purity, whereas FastDNA® SPIN Kit provides a higher DNA yield, but lower purity because the product is mixed with high levels of proteins.[24] To optimise extraction quality, bead beating combined with lyticase digestion is recommended to enhance DNA release from fungal cell wall lysis,[24,25] and column-based DNA purification techniques can enhance the end-product yield and purity.[24,25]

After DNA extraction from faecal samples, the sequencing platform is another factor that affects mycobiota evaluation. Direct sequencing of fungal DNA has been the main method for characterising the mycobiota. Common sequencing targets include the 18S small-subunit ribosomal DNA (rDNA) and 28S large-subunit rDNA. Traditional metagenomic sequencing lacks resolution in evaluating the mycobiota composition in the human gut[26] due to the dominance of bacterial community and that gut fungi only constitute approximately 0.1% of the gut microbial communities.[16–18,27] Deeper metagenomic sequencing and the extension of fungal databases can enhance sequencing output and fungal species identification.[28] Due to current limitations of metagenomic sequencing, a more objective metabar coding technique known as ITS (internal transcribed spacer) sequencing is routinely implemented to specifically target the mycobiota.[29,30] However, mis-attribution of sequences and classification of sexual and asexual forms of the same fungus are current challenges in characterising fungal populations using next-generation DNA sequencing technology. Unlike bacterial 16S rDNA sequences whereby large databases have been developed, the sequences for fungi available in the NCBI GenBank database are far from complete and estimates suggest that only less than 1% of fungal species are currently represented.[31]

The mammalian colon harbours the highest concentration of fungal organisms.[22] The most abundant fungal genera in healthy individuals are Candida, Saccharomyces and Cladosporium.[22,32] Intestinal or faecal mycobiota appears to be less stable and more susceptible to episodic fluctuations than bacterial microbiota.[32] More recently, fungi composition in healthy intestinal mucosa were elucidated and the phyla Ascomycota and Basidiomycota were shown to be the most abundant taxa in the mucosal samples of healthy individuals.[33] The classes Saccharomycetes and Tremellomycetes are dominant in the phyla Ascomycota and Basidiomycota respectively. Further classification at a lower taxonomic level showed that these two classes could be subdivided into the Candida, Debaryomyces, Saccharomyces, Malassezia, Sporobolomyces, Trichosporon, Wallemia genera, along with a smaller proportion of unidentified Filobasidiaceae and Xylariales.[33]

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